Scientists Capture First Images of How X-rays Damage
Proteins

UPTON, NY-While attempting to "photograph" the chemical
reactions of an important enzyme of the nervous system, an international
team of scientists found that the "flash" they were
using - a high-intensity X-ray beam - was systematically destroying
their target. The resulting "movie" of molecular images
is the first-ever direct observation of how proteins break apart
when exposed to high-energy X-rays.

"The observation was stunning," said collaborator
Joel Sussman, formerly on the staff and now a visiting biologist
at the U.S. Department of Energy's Brookhaven National Laboratory,
where some of this research took place. Stunning because, previously,
scientists believed radiation damage was nonspecific, or random.
But the Brookhaven work and studies with other enzymes elsewhere
confirm that the X-rays selectively break particular chemical
bonds.

"It looks like we are seeing 'weak points' in protein
structures that are particularly sensitive to ionizing radiation,"
says Sussman, who is now principally affiliated with the Weizmann
Institute of Science in Israel. Certain disulfide bonds, which
often bridge protein chains, and carboxyl acids, such as those
found at an enzyme's "active site" where reactions take
place, seem particularly vulnerable. Understanding these weak
links may lead to improved methods of preventing high-dose radiation
damage.

Organisms are constantly exposed to radiation, mainly from
natural sources, such as sunlight and cosmic rays, as well as
man-made sources such as diagnostic X-rays. "The ability
to visualize the specific damage caused by radiation at a 'test-tube'
level offers an important diagnostic tool for developing pharmacological
means to protect against radiation damage," says Israel Silman,
also a guest scientist at BNL's Biology Department. The Weizmann
team and European collaborators, together with Brookhaven scientists,
plan to examine the anti-radiation potential of various substances
that might be used to offer general protection or in an emergency.

The findings of X-ray-induced damage, published in the January
18, 2000, issue of the Proceedings of the National Academy of
Science, will also have implications for the use of X-ray techniques
to decipher molecular structures. In X-ray crystallography, scientists
bombard crystalline samples of proteins with high-intensity X-rays
and, by analyzing how the rays diffract, or bend, they work backward
to decipher the protein's molecular structure. This common technique
is an important research topic at Brookhaven's National Synchrotron
Light Source (NSLS), where some of the research described in the
paper was carried out.

Scientists typically perform these experiments at very cold
(cryogenic) temperatures to minimize the damage caused by X-rays.
But the current research, conducted at cryogenic temperatures,
shows that these techniques do not completely prevent the introduction
of inadvertent changes into experimental samples. It is well known
in science that the mere act of observation may change what you
are trying to observe.

The current findings may lead to changes in procedure to minimize
this effect. For example, "Less intensive radiation may provide
more accurate results," says Gitay Kryger, a Weizmann biologist.

The initial experiments were conducted at the European Synchrotron
Radiation Facility (ESRF) in Grenoble, France, using acetylcholinesterase
taken from the Torpedo fish. The work at Brookhaven used the same
enzyme from humans and fruit flies. Additional experiments on
a different enzyme, lysozyme, from hen egg white, were performed
at the ESRF. Additional collaborators included scientists from
Holland's Bijvoet Center for Biomolecular Research, and the European
Molecular Biology Laboratory Outstation in Grenoble, France.

The study was funded by the U.S. Army Medical and Materiel
Command, The European Union 4th Framework Program in Biotechnology,
the Kimmelman Center for Biomolecular Structure and Assembly (Rehovot,
Israel), and the Dana Foundation. The U.S. Department of Energy
funds operations at Brookhaven's NSLS.

The U.S. Department of Energy's Brookhaven National Laboratory
creates and operates major facilities available to university,
industrial and government personnel for basic and applied research
in the physical, biomedical and environmental sciences, and in
selected energy technologies. The Laboratory is operated by Brookhaven
Science Associates, a not-for-profit research management company,
under contract with the U.S. Department of Energy.

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To view images that show the breakage of proteins
exposed to synchrotron X-rays, see the Weizmann Institute of Science
press release at: